Bioengineered Models of Intramural Uterine Fibroids: A Dual-Platform Approach to Study Molecular Transport Barriers and Biochemical Interactions - PROJECT SUMMARY Uterine fibroids, also known as leiomyoma, affect 70-80% of women in the United States of reproductive age and carry significant health and economic burdens. Fibroids are benign fibroproliferative tumors characterized by excessive extracellular matrix (ECM) deposition and remodeling that alters the biophysical and biochemical microenvironment, cell interactions, and access to signaling molecules (i.e., hormones, growth factors) and treatments. Yet, knowledge gaps remain regarding how these factors influence fibroid behavior, and there are limited preclinical in vitro models for uterine fibroids that recapitulate 3D tissue architecture, cell interactions, and relevant microenvironmental features. There is a significant need for improved in vitro models for uterine fibroids with pathophysiologically relevant cell interactions, tunable biophysical properties, and biochemical signaling to effectively study fibroid biology and advance therapeutic development. The proposal objective is to leverage two novel in vitro models of uterine fibroids—one with tunable transport properties and one representing intramural uterine fibroids (fibroids surrounded by myometrium)—to study how altered signaling from sex hormones (estrogen, progesterone), obesity, and therapeutics affect fibroid growth. This work supports the long-term goal of developing predictive in vitro models of uterine fibroids—across all subtypes—to study fibroid initiation, growth, persistence, and recurrence. Aims will test the central hypothesis that fibrotic biophysical properties within the fibroid microenvironment and fibroid-myometrium interactions contribute to fibroid growth and treatment response by reducing molecular transport and modulating the activity of sex hormones, obesity-related factors, and therapeutics. The objective will be accomplished by two Specific Aims: Aim 1 – Characterize fibroid cell responses to molecular transport barriers using a novel tunable collagen-alginate interpenetrating polymer network (IPN) model and Aim 2 – Establish a self- assembled myometrial tissue ring model with embedded fibroid spheroids for evaluating fibroid cell responses to hormones, obesity, and treatment. In Aim 1, we will change fabrication parameters for type I collagen-alginate IPN hydrogels to tune stiffness to that of myometrium and fibroid tissue and reduce molecular transport. A tissue model with a fibroid-tissue sphere embedded within an IPN hydrogel will then be used to evaluate how reduced molecular transport of bioactive molecules of varying size influences functional outcomes, including proliferation, metabolic, activity, and apoptosis, using high throughput and content multiplexed analysis. In Aim 2, self-assembled uterine smooth muscle tissue rings with embedded uterine fibroid spheroids (fibroid cells, fibroblasts) will undergo separate exposures: 1) estrogen and progesterone hormone at concentrations representing a normal menstrual cycle (luteal and follicular phase) and pregnancy (three trimesters); 2) conditioned medium from lean and obese adipocytes; 3) clinically relevant dose of Leuprolide—a gonadotropin-releasing hormone agonist that suppresses estrogen and progesterone production. Models will be evaluated on fibroid growth and related processes, primarily proliferation, apoptosis, and ECM production. Collectively, these studies will provide new insights into how the fibroid microenvironment influences fibroid growth and treatment response, while establishing innovative in vitro models that serve as platforms for future mechanistic and therapeutic investigation.
